The ability of cells to adhere to one another plays a critical role in development, normal physiology, and disease processes such as inflammation. This ability is mediated by adhesion molecules, generally glycoproteins, expressed on cell membranes. Often, an adhesion molecule on one cell type will bind to another adhesion molecule expressed on a different cell type, forming a receptor counter-receptor pair. Three important classes of adhesion molecules are the integrins, selecting, and immunoglobulin (Ig) superfamily members (see Springer, Nature 346:425 (1990); Osborn, Cell 62:3 (1990); Hynes, Cell 69:11 (1992), all of which are incorporated herein by reference in their entirety for all purposes). These molecules are especially vital to the interaction of leukocytes and platelets with themselves and with the extracellular matrix and vascular endothelium.
P-selectin, also known as CD62, granule membrane protein-140 (GMP-140)/platelet activation-dependent granule external membrane (PADGEM)/LECCAM-3 is a specialized cell-surface receptor on vascular endothelial cells and platelets that is involved in the recognition of various circulating cells. P-selectin is a surface glycoprotein with a lectin-like domain, a region with homology to epidermal growth factor, and a region with homology to complement regulatory proteins (see McEver, Blood Cells 16:73-83 (1990)). The structure of P-selectin is similar to that of two other vascular cell surface receptors, endothelial leukocyte adhesion molecule (ELAM-1) and lymphocyte homing receptor (LHR). The term "selectin" has been suggested for this general class of receptors, because of their lectin-like domain and the selective nature of their adhesive functions.
P-selectin is present on the surface of platelets and endothelial cells in response to a variety of stimuli, where it mediates platelet-leukocyte and endothelium-leukocyte interactions. By contrast, ELAM-1 is expressed only on endothelial cells and LHR is expressed on a variety of leukocytes in endothelial venules of peripheral lymph nodes. Expression of P-selectin is inducible and it is not expressed on unactivated endothelial cells or platelets. P-selectin expression does not require de novo synthesis since it is stored in secretory granules (or Weibel-Palade bodies) in both platelets and endothelial cells. Thus, within minutes of activation of either cell type by thrombin, histamine, or phorbol esters or other humoral factors, P-selectin is rapidly redistributed to the surface of the cell where it is accessible for binding to other cells.
A number of mouse antibodies to P-selectin have been reported (see, e.g., U.S. Pat. No. 4,783,330, WO 91/06632, PCT Application No. FR90/00565, McEver et al., J. Biol. Chem. 259:9799-9804 (1984)); Geng et al., Nature 343:757-760; Parmentier et al., Blood 77:1734-1739 (1991)). An additional antibody to P-selectin has been described in presentations at the Scripps Institute, La Jolla Calif. and at a Keystone Colloquium held Jan. 16, 1992. Certain of these antibodies that block P-selectin mediated adhesion are Ca.sup.++ dependent.
In vitro experiments employing antibodies to P-selectin have provided a limited understanding of the ligand specificity of this receptor. It has been reported that P-selectin binds neutrophils, monocytes and certain carcinoma cells (Bevilacqua et al., Proc. Natl. Acad. Sci. USA 84:9238-9242 (1987); Geng et al., Nature 343:757-760 (1990); Larsen et al., Cell 63:467-474 (1990); and Polley et al. It has been also been reported that P-selectin recognizes Le.sup.x as a high-affinity ligand (Larsen et al., Cell 63:467-474 (1990)). However, subsequent experiments showed that SLe.sup.x -containing oligosaccharide is in fact a more potent inhibitor of P-selectin mediated adhesion than Le.sup.x (Polley et al., Proc. Natl. Acad. Sci. USA 88:6224-6228 (1991)).
The participation of P-selectin in intercellular adhesion in vitro suggests that like other adhesion molecules, P-selectin might participate in cellular interactions that contribute to inflammatory diseases in vivo. However, the specific inflammatory diseases in which P-selectin might play a major role have not yet been elucidated. Similarly, the epitope-specificity of agents effective to abort particular P-selectin-mediated inflammatory diseases is also unclear. Of the known antibodies to P-selectin, some bind to P-selectin but do not block P-selectin mediated intercellular interactions. Other known antibodies have the capacity to block P-selectin-mediated intercellular binding only in the presence of high concentrations of Ca.sup.2+, a condition that may not always exist in vivo. The absence of high concentrations of Ca.sup.2+ in vivo would render these antibodies ineffective as therapeutic agents. Moreover, all antibodies to P-selectin produced to-date have been of murine origin and might therefore induce a human-antimouse antibody response (HAMA) in clinical use.
Based on the foregoing it is apparent that a need exists for novel agents targeted against P-selectin that have the appropriate epitope-specificity to abort intercellular interactions leading to inflammatory diseases in vivo. Ideally such agents would not induce a HAMA response in treatment of humans. The present invention fulfills this and other needs.